POS0234 P.E148Q POTENTIATES PYRIN INFLAMMASOME ACTIVATION
Authors: T. Reygaerts, P. Laohamonthonkul, K. Hrovat-Schaale, F. Moghaddas, P. Baker, P. Gray, S. Masters
Journal: Annals of the Rheumatic Diseases (2023)

<h3>Background</h3> The p.E148Q variant in pyrin is present in different populations at a frequency up to 29%[1], and has been associated with inflammatory diseases including vasculitis and familial Mediterranean fever (FMF)[2,3]. The pathogenicity of p.E148Q in FMF is unclear, highly debated in the literature especially when observed <i>in cis or in trans</i> to a single, typically recessive, mutation. If it could modify the expressivity of a pathogenic allele and acts as a risk factor for diseases is not known. <h3>Objectives</h3> We Interrogated the Australian Autoinflammatory Disease RegistrY (AADRY) to find candidate inheritance patterns for the p.E148Q variant in pyrin. We then performed functional validation to determine whether p.E148Q increases the ability of pyrin to form an active inflammasome complex in cell lines. Finaly, the ability of p.E148Q to increase the effect of a known pathogenic mutation in <i>cis</i> or <i>trans</i> was assessed. <h3>Methods</h3> The allele p.E148Q in patients with auto-inflammatory diseases and their families in AADRY was assessed. Different pyrin variant combinations were tested in HEK293T cells stably expressing the adaptor protein apoptosis-associated speck-like (ASC) which were analysed by flow cytometry to visualise inflammasome formation, with and without stimulation by <i>Clostridium difficile</i> toxin B (TcdB). Inflammasome dependent cytokine secretion was also quantified by ELISA of supernatants from THP-1 cells transduced with lentiviral expression vectors. <h3>Results</h3> In AADRY, we observed the p.E148Q allele in individuals with autoinflammatory diseases alone or in conjunction with other pyrin variants. Two FMF families harbored the allele p.M694I-E148Q <i>in cis</i> with dominant heritability. In vitro, p.E148Q pyrin could spontaneously potentiate inflammasome formation, with increased IL-1β and IL-18 secretion. p.E148Q <i>in cis</i> to classical FMF mutations provided significant potentiation of inflammasome formation. <h3>Conclusion</h3> The p.E148Q variant in pyrin potentiates inflammasome activation in vitro. <i>In cis</i>, this effect is additive to known pathogenic FMF mutations. These observations may help to explain the association of p.E148Q with FMF and other inflammatory diseases. <h3>References</h3> [1]K. J. Karczewski, L. C. Francioli, G. Tiao, B. B. Cummings, J. Alföldi, Q. Wang, et al. The mutational constraint spectrum quantified from variation in 141,456 humans. Nature 2020 Vol. 581 Issue 7809 Pages 434-443. [2]D. Cattan, MEFV mutation carriers and diseases other than familial Mediterranean fever: proved and non-proved associations; putative biological advantage, Curr Drug Targets Inflamm Allergy 2005 Vol. 4 Issue 1 Pages 105-12. [3]I. Touitou, The spectrum of Familial Mediterranean Fever (FMF) mutations, Eur J Hum Genet 2001 Vol. 9 Issue 7 Pages 473-83. <h3>Acknowledgements</h3> We would like first to thank the patients and families involved in AADRY. We are grateful to all members of the Masters laboratory for their support. We thank the WEHI flowcytometry facility (Simon Monard) for excellent technical support. <h3>Disclosure of Interests</h3> Thomas Reygaerts: None declared, Pawat Laohamonthonkul: None declared, Katja Hrovat-Schaale: None declared, Fiona Moghaddas: None declared, Paul Baker: None declared, Paul Gray: None declared, Seth Masters Consultant of: Scientific Advisor for Odyssey Therapeutics., Employee of: VP of Discovery Biology for NRG Therapeutics.